Molecular epidemiology and antibiotic susceptibility pattern of Acinetobacter baumannii isolated from children in a Turkish university hospital

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1 The Turkish Journal of Pediatrics 2014; 56: Original Molecular epidemiology and antibiotic susceptibility pattern of Acinetobacter baumannii isolated from children in a Turkish university hospital Özlem Özgür Gündeşlioğlu 1, Tülin Güven Gökmen 2, Özden Özgür Horoz 3, Necmi Aksaray 1, Fatih Köksal 2, Akgün Yaman 2, Rıza Dinçer Yıldızdaş 3, Emre Alhan 1, Emine Kocabaş 1, Derya Alabaz 1 Division of 1 Pediatric Infectious Diseases and 3 Pediatric Intensive Care Unit, Department of Pediatrics, and 2 Department of Clinical Microbiology, Cukurova University Faculty of Medicine, Adana, Turkey ozlemozgur1978@yahoo.com SUMMARY: Gündeşlioğlu ÖÖ, Gökmen TG, Horoz ÖÖ, Aksaray N, Köksal F, Yaman A, Yıldızdaş RD, Alhan E, Kocabaş E, Alabaz D. Molecular epidemiology and antibiotic susceptibility pattern of Acinetobacter baumannii isolated from children in a Turkish university hospital. Turk J Pediatr 2014; 56: The aim of the present study is to investigate the types of healthcareassociated infections (HC-AIs) caused by Acinetobacter baumannii and the related antibiotic susceptibility patterns as well as the genotypic characteristics of the Acinetobacter baumannii isolates from our center. Sixty-nine Acinetobacter baumannii isolates originating from various samples collected from 69 pediatric patients during their hospital stays were included in the study. The types of healthcare-associated infections caused by these isolates were evaluated, and the antibiotic susceptibility pattern and the genotypic characteristics of the isolates were determined using the pulsed-field gel electrophoresis (PFGE) method. Fifty of the 69 children were observed to have HC-AIs, and 19 children had Acinetobacter baumannii colonization. Healthcare-associated pneumonia (58%) was the most common type of these infections. The rate of carbapenem resistance was found as 91.3%, while tigecycline resistance was found as 18.84%. No colistin resistance was observed in any of the isolates. A total of 10 groups, comprising eight major and two minor groups, were determined using the pulsed-field gel electrophoresis method. Acinetobacter baumannii isolates are the leading cause of healthcare-associated infections, and they show high rates of multidrug antibiotic resistance. Molecular epidemiological evaluation using PFGE plays an important role in preventing healthcare-associated infections. Key words: Acinetobacter baumanni, pulsed-field gel electrophoresis, healthcareassociated infections, antibiotic resistance, children. Members of the genus Acinetobacter have been increasingly responsible for healthcareassociated infections, especially in intensive care units, in recent years 1. The most frequently isolated type of Acinetobacter from clinical samples, Acinetobacter baumannii (A. baumannii), has a low virulence and usually leads to opportunistic infections, including primarily ventilator-associated pneumonia and bacteremia, urinary system infections, meningitis and skin and soft tissue infections in patients admitted to intensive care units 2-4. A. baumannii has developed resistance to various types of antibiotics, including carbapenems, and multi-resistant epidemics of A. baumannii have recently been reported 5-8. In order to control HC-AIs and epidemics, it is of vital importance to know the origins and transmission routes of the isolates. In the past, phenotypic methods were used in order to compare the similarities and differences among isolates originating from different sources in healthcare-associated infections or epidemics. In recent years, this approach has changed in parallel to the progress in the molecular typing methods based on deoxyribonucleic acid

2 Volume 56 Number 4 Acinetobacter baumannii Infections in Children 361 (DNA). Today, pulsed-field gel electrophoresis (PFGE) is accepted as the gold standard among the frequently used DNA-based molecular typing methods 9. Material and Methods The present study was conducted at the Cukurova University Medical Faculty Hospital between February 2010 and October For the purposes of the study, approval was granted by the Cukurova University Medical Faculty Ethics Committee. The study had a prospective design based on the A. baumannii strains isolated from various clinical samples routinely sent to the microbiology laboratory from pediatric patients admitted to the Cukurova University Medical Faculty Hospital. In cases where more than one A. baumannii isolate was derived from a single patient, only one of the isolates was included in the study. The epidemiologic, clinical and demographic characteristics of the patients were recorded. The clinical data gathered included the patient s age, gender, duration of hospital stay, sites of infection, time from admission until collection of the sample, and any comorbidities or major risk factors (e.g., intensive care unit stay, intravenous catheterization, mechanical ventilation). The definitions of healthcare-associated infections and colonization were based on the U.S. Centers for Disease Control and Prevention (CDC) criteria 10. Assessment of Antibiotic Susceptibility Antibiotic susceptibility was tested using the Vitek-2 Compact automated system. The MIC values of the antibiotics are presented in Table I 11. Intermediate-level susceptible strains were also accepted as resistant. The Pulsed-Field Gel Electrophoresis Method Plug preparation, lysis, cell washing, restriction digestion and electrophoresis were performed as has been previously described 12. For the electrophoresis, a CHEF-DR II (Bio-Rad Laboratories, Nazareth, Belgium) device was used. The band profiles were analyzed using GelCompar II software (version 5.0; Applied Maths, Sint-Martens-Latem, Belgium). The band and profile tolerance was taken as 1.5% in the calculation of the similarity coefficient. The isolates where the band profiles showed an 80% similarity were evaluated within the same group and designated with capital letters. Subtypes within the same group were indicated with numbers. Results Sixty-nine Acinetobacter baumannii isolates originating from various clinical samples collected from 69 pediatric patients during their hospital stays were included in the study. When the patients were assessed in terms of the infections that developed due to A. baumannii and the risk factors for colonization, all of them were observed to be under treatment with extended-spectrum antibiotics. The risk factors are presented in Table II. In this study, 50 among the 69 isolates (72.74%) were the agents of healthcareassociated infections; the most frequently observed infection (50%) was ventilatorassociated pneumonia. In 19 patients (27.53%), the isolated A. baumannii was accepted as colonization. The diagnoses of the patients are presented in Table III. Antibiotic susceptibility results Among the isolates included in the study, 67 (100%) were observed to be susceptible to colistin. No antibiogram was performed against colistin for the two isolates included in the study. The antibiotic susceptibility of the isolates included in the study is presented in Table IV. Pulsed-Field Gel Electrophoresis Results The pulsed-field gel electrophoresis results indicated 10 different clones (A-J), including eight major groups and two separate isolates, with more than 80% band profile similarity among the 69 isolates. The largest group among these clones was group D, with 23 isolates. Seven subtypes were detected within group D (D1-D7). Group F was the second largest group, with 16 isolates and nine subtypes (F1-F9). Group C included seven isolates and seven subtypes (C1-C7); group E had three isolates and three subtypes (E1-E3); group G comprised five isolates and three subtypes (G1-G3); group H had three isolates and three subtypes (H1-H3); group I included five isolates and four subtypes (I1-I4), and group J comprised five isolates and four subtypes (J1-J4). The dendrogram obtained through the pulsed-field gel electrophoresis analysis is presented in Figure 1.

3 362 Gündeşlioğlu ÖÖ, et al The Turkish Journal of Pediatrics July-August 2014 Fig. 1. Pulsed-field gel electrophoresis dendrogram of the isolates included in the study

4 Volume 56 Number 4 Acinetobacter baumannii Infections in Children 363 Our study has demonstrated that different clones may be present within the same hospital units, while the same clones may be present in different units. The clone with the longest lifespan was in group D, and these isolates were first detected in the pediatric intensive care unit and last in the burn unit. This clone has been demonstrated to have persisted in different services in our hospital between 28 March 2010 and 09 May Also, we have observed that the PFGE type C clone persisted in the pediatric intensive care unit in the period between 16 July 2010 and 16 March Discussion Healthcare-associated infections caused by resistant microorganisms and especially by carbapenem-resistant A. baumannii strains have become an important cause of morbidity and mortality in hospitals in recent years 13. According to the data from the infection control committee of our hospital, A. baumannii was the most frequently observed microorganism, with a ratio of 16.34% in terms of the distribution according to the species of the agents isolated from the infections originating in the Intensive Care Units of our hospital during our study. A prolonged hospital stay, surgical intervention, wounds, previous infections and extendedspectrum antibiotic use, central venous or urinary system catheterization, an intensive care stay, a burn unit stay, parenteral nutrition, mechanical ventilation and shortcomings in the infection control program are the risk factors for healthcare-associated infections caused by Acinetobacter baumanni In our study, the most common risk factor was extendedspectrum antibiotic use, which was observed in all of the patients. The other risk factors found in our patients were (in descending order) use of a central venous catheter, mechanical ventilation, an intensive care stay, surgical intervention, total parenteral nutrition, urinary system catheterization, presence of wounds disrupting the integrity of the skin, and a burn unit stay. Healthcare-associated infections due to A. baumannii may involve any site. In most institutions, A. baumannii is an increasingly important cause of healthcare-associated pneumonia in the intensive care unit, particularly in patients with ventilator-associated pneumonia, representing between 5% and 10% of the cases 9,17. A. baumannii may lead to healthcare-associated bacteremia and infections of the skin and soft tissue, genitourinary system or intracranial system 2,9,18. Among the 69 A. baumannii isolates in our study, 19 (27.53%) were defined as colonizations. More than half (58%) of the 50 isolates thought to have Table I. The Antibiotics and MIC Values 11 Antibiotic MIC Interpretation Ampicillin/Sulbactam 32 Resistant Piperacillin 128 Resistant Piperacillin/tazobactam 128 Resistant Ceftazidime 64 Resistant Cefoperazone/sulbactam 32 Intermediate-level susceptible Cefepime 64 Resistant Imipenem 16 Resistant Meropenem 16 Resistant Amikacin 32 Intermediate-level susceptible Gentamicin 1 Susceptible Netilmicin 8 Susceptible Ciprofloxacin 4 Resistant Levofloxacin 4 Intermediate-level susceptible Tetracycline 16 Resistant Tigecycline 2 Susceptible Colistin 0.5 Susceptible Trimethoprim/Sulfamethoxazole 320 Resistant

5 364 Gündeşlioğlu ÖÖ, et al The Turkish Journal of Pediatrics July-August 2014 Table II. Frequency of the Risk Factors That May Lead to Acinetobacter baumannii Infections in the Patients Included in the Study Risk Factors Number of patients % Underlying disease Surgical intervention Wounds on the body Treatment with extended-spectrum antibiotics Central venous catheter Urinary system catheter Intensive care stay Burn unit stay Total parenteral nutrition Mechanical ventilation caused healthcare-associated infections were assessed to be agents associated with ventilatorassociated pneumonia. Although this ratio is in correlation with the studies reporting that A. baumannii most often leads to healthcareassociated pneumonia, it seems to be higher than the ratios reported in the literature. This higher ratio observed in our patients may be explained by the fact that the majority of our patients were mechanically ventilated or in the ICU. The other infections observed in our patients related to A. baumannii were (in descending order) central line-associated bloodstream infections, laboratory-evidenced bacteremia, meningitis, surgical site infections, burn wounds, permanent catheter-related bloodstream infections, infected decubitus ulcers, urinary system infections and rectal abscesses. As in rest of the world, the A.baumannii strains were multi-antibiotic resistant. In a multicenter study conducted in Europe in 1999, susceptibility among the Acinetobacter strains isolated from intensive care units was observed to be greatest to imipenem 19. Similarly, according to the industry-supported MYSTIC (Meropenem Yearly Susceptibility Test Information Collection) report, which evaluated antibiotic resistance in 490 Acinetobacter baumannii isolates collected between 1997 and 2000 from 37 centers in 11 European countries, imipenem and meropenem were the two most effective antibiotics against A. baumanni, although resistance was also observed in ratios of 16% and 14%, respectively 20. However, according to the MYSTIC report dated 2006, the resistance against these two antibiotics had shown a significant increase and reached 40% 21. Studies supporting these data and observing carbapenem resistance rates Table III. Types of Infection Caused by the Strains Included in the Study, and Colonization Rates Number of patients (n=69) % Type of healthcare-associated infection (n=50; 72.47%) Ventilator-associated pneumonia Catheter-associated bloodstream infection 6 12 Laboratory-confirmed bloodstream infection 4 8 Meningitis or ventriculitis 3 6 Incisional surgical site infection 3 6 Burn wound infection 2 4 Decubitus ulcer infection 1 2 Catheter-related urinary tract infection 1 2 Gastrointestinal tract infection (rectal abscess) 1 2 Colonization 19 (27.53%)

6 Volume 56 Number 4 Acinetobacter baumannii Infections in Children 365 Antibiotic Table IV. Antibiotic Susceptibility of the Isolates Included in the Study Susceptible isolate Resistant isolate n % n % Amikacin Ampicillin/Sulbactam Ciprofloxacin Colistin Ceftriaxone Cefepime Gentamicin Imipenem Meropenem Levofloxacin Piperacillin Piperacillin/tazobactam Ceftazidime Cefoperazone/sulbactam Ticarcillin Trimethoprim/Sulfamethoxazole Tetracycline Tigecycline Tobramycin up to 100% have been reported from various geographical regions around the world 22,23,24. Studies from Turkey have reported carbapenem resistance rates of 78% for imipenem and 71% for meropenem 25,26. Parallel to these data, our study has also pointed out high rates of imipenem and meropenem resistance, both reaching 91.3%. The carbapenem resistance we observed in the strains isolated from our patients seems to be higher than the ratios reported by other centers in our country. This high resistance against carbapenems may be explained by a number of factors. Firstly, the majority of our subjects were ICU patients. Secondly, all the patients had a history of treatment with extended-spectrum antibiotics. And finally, they were frequently hospitalized patients due to other underlying diseases. The high carbapenem resistance observed in Acinetobacter baumannii has led to a search for new treatment options. Certain studies have reported a % susceptibility to colistin On the other hand, some studies have reported no resistance to colistin 25,31,32, while a study from our country has reported a 12.1% resistance 33. In different studies, the resistance rates to tigecycline varied from 5 to 78% In a study that evaluated 492 carbapenem-resistant Acinetobacter baumannii isolates from various regions in the world, including Turkey, susceptibility to tigecycline and minocycline was reported to be over 80%, although the susceptibility to other antibiotics was low (38%). As observed in these studies, the antibiotics with the lowest resistance rates for the treatment of multiresistant Acinetobacter baumannii infections are colistin and tigecycline In line with the literature, our study has also shown high resistance rates to other antibiotics, while the resistance to tigecycline was only 18.84%, and no colistin resistance was observed in any of the strains. Numerous publications have reported the phenotypic and genotypic features of the strains isolated during epidemics. The most common method currently used for the genotyping of A. baumannii is pulsed-field gel electrophoresis (PFGE). Although certain clones are present with widespread dissemination, isolates of A. baumannii from hospitals in the same country or even from a single hospital may show significant genetic diversity 12, 24,31,39-45.

7 366 Gündeşlioğlu ÖÖ, et al The Turkish Journal of Pediatrics July-August 2014 Our study also revealed that many different PFGE genotypes were present during the 1.5 year period. We have determined that clonally related strains can survive for a long time in our hospital and cause healthcareassociated infections at different times. The PFGE molecular typing of the Acinetobacter baumannii strains isolated during the 1.5 year period of our study has revealed that the same isolates were found in different units and some isolates persisted in the units for long periods. These findings point to the transmission of infections between the units and a lack of efficient measures for infection control. In conclusion, it can be claimed that A. baumannii plays an important role in healthcare-associated infections, and high antibiotic resistance is observed in this agent. The increasing costs of the morbidity, mortality and treatment related to healthcare-associated infections call for the implementation of infection control strategies. For every center, studying the patient profile, the microorganisms composing the hospital flora together with their resistance patterns, and the distribution and frequency of healthcareassociated infections in each unit will help in developing the right strategies. Pulsed field gel electrophoresis, which is the gold standard among molecular typing methods, should be employed in long-term surveillance in order to prevent healthcare-associated infections, determine the transmission sources and routes of microorganisms, and monitor the spread of these strains throughout the hospital. REFERENCES 1. Fournier PE, Richet H. The epidemiology and control of Acinetobacter baumannii in health care facilities. Clin Infect Dis 2006; 42: Allen DM, Hartman BJ. Acinetobacter species. In: Mandell GL, Bennett JE, Dolin R (eds). Principles and Practice of Infectious Diseases (7th ed). Philadelphia: Elsevier/Churchill Livingstone; 2010: Towner KJ. Acinetobacter: an old friend, but a new enemy. J Hosp Infect 2009; 73: Garnacho-Montero J, Ortiz-Leyba C, Fernández- Hinojosa, et al. Acinetobacter baumannii ventilatorassociated pneumonia: epidemiological and clinical findings. Intensive Care Med 2005; 31: Gordon NC, Wareham DW. A review of clinical and microbiological outcomes following treat ment of infections involving multidrug-resistant Acinetobacter baumannii with tigecycline. J Antimicrob Chemother 2009; 63: Poirel L, Nordmann P. Carbapenem resistance in Acinetobacter baumannii: mechanisms and epidemiology. Clin Microbiol Infect 2006; 12: Peleg AY, Potoski BA, Rea R, et al. Acinetobacter baumannii bloodstream infection while receiving tigecycline: a cautionary report. J Antimicrob Chemother 2007; 59: Zarrilli R, Giannouli M, Tomasone F, Triassi M, Tsakris A. Carbapenem resistance in Acinetobacter baumannii: the molecular epidemic features of an emerging problem in health care facilities. J Infect Dev Ctries 2009; 3: Peleg AY, Seifert H, Paterson DL. Acinetobacter baumannii: emergence of a successful pathogen. Clin Microbiol Rev 2008; 21: Horan TC, Andrus M, Dudeck MA. CDC/NHSN surveillance definition of healthcare-associated infection and criteria for specific types of infections in the acute care setting. Am J Infect Control 2008; 36: Clinical and Laboratory Standards Institute. Performance Standards for Antimicrobial Susceptibility Testing; Twentieth Informational Supplement, M100-S20-U. Wayne, PA: Clinical and Laboratory Standards Institute; 2010: Seifert H, Dolzani L, Bressan R, et al. Standardization and interlaboratory reproducibility assessment of pulsed-field gel electrophoresis-generated fingerprints of Acinetobacter baumannii. J Clin Microbiol 2005; 43: Giamarellou H, Poulakou G. Multidrug-resistant Gramnegative infections: what are the treatment options? Drugs 2009; 69: Anstey NM, Currie BJ, Withnall KM. Communityacquired Acinetobacter pneumonia in the Northern Territory of Australia. Clin Infect Dis 1992; 14: Lortholary O, Fagon JY, Hoi AB, et al. Nosocomial acquisition of multiresistant Acinetobacter baumannii: risk factors and prognosis. Clin Infect Dis 1995; 20: Donskey CJ. Antibiotic regimens and intestinal colonization with antibiotic-resistant gram-negative bacilli. Clin Infect Dis 2006; 43 (Suppl 2:) S McDonald LC, Banerjee SN, Jarvis WR. Seasonal variation of Acinetobacter infections: Nosocomial Infections Surveillance System. Clin Infect Dis 1999; 29: Manchanda V, Sanchaita S, Singh NP. Multidrug resistant Acinetobacter. J Glob Infect Dis 2010; 2: Hanberger H, Garcia-Rodriguez JA, Gobernado M, Goossens H, Nilsson LE, Struelens MJ. Antibiotic susceptibility among aerobic gram-negative bacilli in intensive care units in 5 European countries. French and Portuguese ICU Study Groups. JAMA 1999; 281: Turner PJ, Greenhalgh JM; MYSTIC Study Group (Europe) The activity of meropenem and comparators against Acinetobacter strains isolated from European hospitals, Clin Microbiol Infect 2003; 9:

8 Volume 56 Number 4 Acinetobacter baumannii Infections in Children Turner PJ. Meropenem activity against European isolates: report on the MYSTIC (Meropenem Yearly Susceptibility Test Information Collection) 2006 results. Diagn Microbiol Infect Dis 2008; 60: Cisneros JM, Rodríguez-Baño J. Nosocomial bacteremia due to Acinetobacter baumannii: epidemiology, clinical features and treatment. Clin Microbiol Infect 2002; 8: Trottier V, Segura PG, Namias N, King D, Pizano LR, Schulman CI. Outcomes of Acinetobacter baumannii infection in critically ill burned patients. J Burn Care Res 2007; 28: Irfan S, Turton JF, Mehraj J, et al. Molecular and epidemiological characterisation of clinical isolates of carbapenem-resistant Acinetobacter baumannii from public and private sector intensive care units in Karachi, Pakistan. J Hosp Infect 2011; 78: Kurtoğlu MG, Opuş A, Kaya M, Keşli R, Güzelant A, Yüksekkaya Ş. Bir eğitim ve araştırma hastanesinde klinik örneklerden izole edilen Acinetobacter baumannii suşlarında antibakteriyel direnç ( ). Ankem Derg 2011; 25: Bacakoğlu F, Korkmaz Ekren P, Taşbakan MS, et al. Solunumsal yoğun bakım ünitesinde çoklu antibiyotik dirençli Acinetobacter baumannii enfeksiyonu. Mikrobiyol Bült 2009; 43: Henwood CJ, Gatward T, Warner M, et al. Antibiotic resistance among clinical isolates of Acinetobacter in the UK, and in vitro evaluation of tigecycline (GAR- 936). J Antimicrob Chemother 2002; 49: Luyt CE, Combes A, Nieszkowska A, Trouillet JL, Chastre J. Aerosolized antibiotics to treat ventilatorassociated pneumonia. Curr Opin Infect Dis 2009; 22: Gales AC, Jones RN, Sader HS. Global assessment of the antimicrobial activity of polymyxin B against clinical isolates of Gram-negative bacilli: report from the SENTRY antimicrobial surveillance programme ( ). Clin Microbiol Infect 2006; 12: Souli M, Kontopidou FV, Koratzanis E, et al. İn vitro activity of tigecycline against multiple-drugresistant, including pan-resistant, gram-negative and gram-positive clinical isolates from Greek hospitals. Antimicrob Agents Chemother 2006; 50: Schimith Bier KE, Luiz SO, Scheffer CM, et al. Temporal evolution of carbapenem-resistant Acinetobacter baumannii in Curitiba, southern Brazil. Am J Infect Control 2010; 38: Falagas ME, Kasiakou SK. Colistin: the revival of polymyxins for the management of multidrug-resistant gram-negative bacterial infections. Clin Infect Dis 2005; 40: Çetin ES, Durmaz R, Tetik T, Otlu B, Kaya S, Çalışkan A. Epidemiologic characterization of nosocomial Acinetobacter baumannii infections in a Turkish university hospital by pulsed-field gel electrophoresis. Am J Infect Control 2009; 37: Peterson LR: A review of tigecycline - the first glycylcycline. Int J Antimicrob Agents 2008; 32 Suppl 4: S Dominguez EA: Single-agent therapy with tigecy cline in the treatment of complicated skin and skin structure and complicated intraabdominal infecti ons. Infect Dis Clin Pract 2009; 17: Seifert H, Stefanik D, Wisplinghoff H. Comparative in vitro activities of tigecycline and 11 other antimicrobial agents against 215 epidemiologically defined multidrugresistant Acinetobacter baumannii isolates. J Antimicrob Chemother 2006; 58: Dizbay M, Altuncekic A, Sezer BE, Ozdemir K, Arman D. Colistin and tigecycline susceptibility among multidrug-resistant Acinetobacter baumannii isolated from ventilator-associated pneumonia. Int J Antimicrob Agents 2008; 32: Higgins PG, Dammhayn C, Hackel M, Seifert H. Global spread of carbapenem-resistant Acinetobacter baumannii. J Antimicrob Chemother 2010; 65: Nemec A, Dijkshoorn L, van der Reijden TJ. Long-term predominance of two pan-european clones among multi-resistant Acinetobacter baumannii strains in the Czech Republic. J Med Microbiol 2004; 53: Abbo A, Navon-Venezia S, Hammer-Muntz O, Krichali T, Siegman-Igra Y, Carmeli Y. Multidrug-resistant Acinetobacter baumannii. Emerg Infect Dis 2005; 11: Zarrilli R, Vitale D, Di Popolo A, et al. A plasmidborne blaoxa-58 gene confers imipenem resistance to Acinetobacter baumannii isolates from a Lebanese hospital. Antimicrob Agents Chemother 2008; 52: Runnegar N, Sidjabat H, Goh HM, Nimmo GR, Schembri MA, Paterson DL. Molecular epidemiology of multidrug-resistant Acinetobacter baumannii in a single institution over a 10-year period. J Clin Microbiol 2010; 48: Dijkshoorn L, Aucken H, Gerner-Smidt P, et al. Comparison of outbreak and nonoutbreak Acinetobacter baumannii strains by genotypic and phenotypic methods. J Clin Microbiol 1996; 34: Di Popolo A, Giannouli M, Triassi M, Brisse S, Zarrilli R. Molecular epidemiological investigation of multidrug-resistant Acinetobacter baumannii strains in four Mediterranean countries with a multilocus sequence typing scheme. Clin Microbiol Infect 2011; 17: Carretto E, Barbarini D, Dijkshoorn L, et al. Widespread carbapenem resistant Acinetobacter baumannii clones in Italian hospitals revealed by a multicenter study. Infect Genet Evol 2011; 11: